We have observed that replicating DNA is more vulnerable to attack by chemical carcinogens than the bulk of the nuclear DNA and that DNA damage occuring in the early part of the S phase is far more effective than damage at other times in producing transformation of cells. We believe that DNA sites replicating during the early S phase were probably the sites in DNA segments that are essential to the transformation process. The vastly higher transformation rates result from the greater likelihood that these DNA sites would be damaged and genetically altered as the result of treatments with carcinogens early in the S phase. The goal of this project is to identify sites in DNA that might play this critical role in transformation. The concept is that DNA replicated early in the S phase should include DNA segments closes to and at replication origins. Alteration of such sites in DNA could offer means whereby the fine regulation of genomic DNA replication could be degraded. Based on observations that DNA replication occurs and starts at sites physically organized and related to the nuclear matrix, we propose a search strategy for such sites based on their presence in both early S DNA libraries and matrix-associated DNA libraries. In addition, studies will be undertaken to assess whether these sites are altered in transformed cells, or even early stage, partially-transformed cells as compared to normal cells.